Integrated molecular diagnostic system for the point-of-care

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DESCRIPTION (provided by applicant): This STTR-AT-NIAID seeks to develop an integrated nucleic acid system based on research done by Catherine Klapperich's laboratory at Boston University. The BU lab-on-a-chip includes a micro solid phase extraction (<SPE) column, flap valves and hydrophobic vents to gate fluid movements through micro channels, and multiple reaction chambers for experimental replicates and control reactions. Dr. Klapperich's laboratory has successfully performed nucleic acid amplifications in the chip using BioHelix's isothermal amplification process. Biohelix's proprietary technology is called helicase-dependent amplification (HDA). It uses DNA helicases to separate DNA strands during exponential amplification at a constant temperature of 65:C. Like the polymerase chain reaction (PCR), HDA assays use a competitive internal control (i.e., a template DNA of known concentration spiked into the raw sample) that can be amplified by the same primers as the analyte, but detected separately; hence allowing us to detect amplification inhibitors in direct clinical samples. The objectives of Phase I are to: 1) modify the BU microfluidic device design to incorporate a lateral flow strip as a means of detecting amplification products using the naked eye; and 2) demonstrate the feasibility of performing integrated tests using the device from aim 1 using CT and NG spiked at 104 in 1 mL urine to establish proof of concept for integrated assays performed in a modified BU chip that includes a lateral flow strip. Our specific aims for Phase II will be to develop a lateral flow reader sub-system, integrate it with the BU instrument to build a pre-commercial looks-like / works-like prototypes, simplify chip design and fabrication to allow for pilot scale manufacturing, and implement a pilot scale manufacturing for the disposable at BioHelix / Quidel. At this stage Qiagen is the most likely manufacturer for the reader. At the conclusion of Phase II, we should be ready for clinical validation of the new assay system. PUBLIC HEALTH RELEVANCE: This Phase I STTR-AT-NIAID project seeks proof-of-concept project will focus on the most abundant sexually transmitted disease (STD) pathogens: Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG). The scientific literature clearly shows that molecular testing is the most sensitive means of detecting CT and NG and the molecular CT/NG high throughput screening market is currently valued at over $300M/year. Moreover, CDC urges STD clinics to test patients with POC tests if health care workers suspect these patients are unlikely to return to the STD clinic to learn the results of the test. Unfortunately, there are no point-of-care (POC) CT NG molecular tests, and existing POC molecular testing systems like the GeneXpert are too costly for use in STD clinics. This proposed project would seek to remedy to this short coming. We propose to develop a low-cost POC molecular diagnostic system using a design developed by Dr. Catherine Klapperich's laboratory at Boston University (BU). Although the current BU disposable can perform our proprietary isothermal amplification reactions, it does not allow for low cost, instrument-free detection of amplification products; i.e., a fluorescence microscope can be used to detect product formation but this is not a commercially viable option. The device we envisage for Phase I will incorporate a lateral flow strip as a means of detecting the presence or absence of nucleic acid amplification products by simple visual inspection. BioHelix has experience in developing molecular tests using lateral flow based detection.